SUMMARY (PROJECT 2) Vaccines represent the best hope for preventing and controlling dengue, yet vaccine development is complicated by the presence of four dengue virus (DENV) serotypes, unbalanced serotype-specific immunity in vaccine recipients, and the possibility of immune-enhanced dengue disease. The three leading dengue vaccine candidates are based on tetravalent live attenuated dengue vaccine (TV-DLAV) formulations, which are currently being tested in clinical trials. For Project 2, we propose to characterize the antibody and B cell responses in people receiving DENVax, a leading TV-DLAV licensed by Takeda Vaccines, Inc. We use DENVax as a model platform to dissect B cell and antibody responses associated with simultaneous exposure to 4 attenuated DENVs. Our group and others have discovered new quaternary structure epitopes on the surface of DENVs that are targeted by strongly neutralizing, serotype-specific antibodies in people exposed to natural DENV infections. We hypothesize that in DENV-nave individuals who receive DENVax (Specific Aim 1), neutralization will be mediated by serotype-specific antibody responses that target these quaternary structure epitopes. Our preliminary studies demonstrate that people exposed to natural secondary DENV infections develop serotype cross-reactive neutralizing antibodies that are qualitatively different from antibodies induced after primary exposure. We propose that similar cross-neutralizing antibodies will be induced when DENV-exposed individuals receive DENVax (Specific Aim 2). Antibody genes will be sequenced to determine if these cross-neutralizing antibodies are derived from somatically mutated memory B cells (MBCs) clones that secrete high affinity antibodies. Currently, there are no established immune correlates of protection from dengue disease. Aim 3 will analyze samples from DENVax Phase 3 efficacy trials to identify antibody responses associated with protection from disease. In this P01, Projects 2 and 3 will analyze the same vaccine samples to obtain a comprehensive view of both the B and T cell response to TV-DLAVs. Moreover, samples collected for Projects 1 and 2 will be matched and characterized using the same assays to directly compare antibody responses to natural DENV infection and vaccination.